Pharyngeal lipse is secreted in the pharynx of man and rat, and it acts in the stomach at pH 5.4 to hydrolyze dietary triacylglycerol to diacylglycerol, monoacylglycerol and fatty acids. These amphipathic products facilitate emulsification of lipid in the stomach before it passes to the small intestines for further digestion and absorption. We have continued our effort to purify this enzyme from rat tongue and have succeeded in obtaining a nearly pure preparation. A protein preparation with lipolytic activity has been obtained from a homogenate of rat lingual serous glands by precipitating from a 100,000 x g supernatant with 60% saturated ammonium sulfate solution, resuspending in aqueous buffer (pH 5.4) and precipitating with cold acetone; this fraction contained 110 units of lipolytic activity per mg. Further purification yielding a specific activity of 700 units per mg was obtained by passing the above protein preparation through the cationic exchange resin SE sephadex G-25. Gel filtration indicates the molecular weight of the enzyme is 48,000 daltons. Protein eluted from a hydrophobic column of ethyl agarose showed one major band, with a molecular weight of 50,000 daltons, on SDS-polyacylamide gel electrophoresis.